Field of the Invention
Embodiments of the invention relate to high density membranes comprising block copolymers and membrane proteins. Further embodiments of the invention include methods for production of such membranes.
Description of the Related Art
Membrane proteins mediate specific and efficient transport of water, ions and solutes across cell membranes. They also serve the cell as sensors that detect environmental conditions, ranging from pH to specific signaling molecules and toxins.
Incorporating such membrane proteins into stable membranes formed by lipids and lipid analogs could provide materials with targeted applications in sensors, enzymatic reactions, drug screening, and even water purification. See Gu, L.; Braha, O.; Conlan, S.; Cheley, S.; Bayley, H. Nature 1999, 398, 686-690; Bayley, H.; Cremer, P. S. Nature 2001, 413, 226-230; Astier, Y.; Bayley, H.; Howorka, S. Curr. Opin. Chem. Biol. 2005, 9, 576-584; Suzuki, H.; Tabata, K.; Kato-Yamada, Y.; Noji, H.; Takeuchi, S. Lab. Chip 2004, 4, 502-505; and Kumar, M.; Grzelakowski, M; Zilles, J.; Clark, M.; Meier, W. Proc. Natl. Acad. Sci. USA 2007, 104, 20719-20724. Block copolymers (BCPs) form membranes that mimic the architecture of lipid bilayers and allow incorporation of functional membrane proteins. See Taubert, A.; Napoli, A.; Meier, W. Curr. Opin. Chem. Biol. 2004, 8, 598-603.
In contrast to lipids, BCPs can be tailored to have the stability and durability associated with polymeric materials and hence are more suitable for the production of membrane protein-based devices and other applications. See Discher, B.; Won, Y.; Ege, D.; Lee, J.; Bates, F.; Discher, D.; Hammer, D. Science 1999, 284, 1143-1146. Furthermore, BCPs can be designed to form membranes with specific physical properties and unique morphologies simply by the choice of the blocks and their lengths or their length ratio. See Discher, D. E.; Eisenberg, A. Science 2002, 297, 967-973; Zhang, L.; Eisenberg, A. Science 1995, 268, 1728-1731. Physical properties that can be engineered include the toughness and permeability of the membrane, as well as its morphology (e.g., micellar, vesicular, cylindrical or planar). BCP end groups can also be modified by molecules such as biotin and 4-formylbenzoate (for recognition and immobilization), methacrylate (for stabilization by crosslinking), fluorescent molecules (for imaging), and even drugs (for drug delivery). See Grzelakowski, M.; Onaca, O.; Rigler, P.; Kumar, M.; Meier, W. Small 2009, 5, 2545-2548; Egli, S.; Nussbaumer, M. G.; Balasubramanian, V.; Chami, M.; Bruns, N.; Palivan, C.; Meier, W. J. Am. Chem. Soc. 2011, 133, 4476-4483; Nardin, C.; Hirt, T.; Leukel, J.; Meier, W. Langmuir 2000, 16, 1035-1041; and Peer, D.; Karp, J. M.; Hong, S.; Farokhzad, O. C.; Margalit, R.; Langer, R. Nature Nanotech. 2007, 2, 751-760.
Recent efforts to insert membrane proteins into BCP membranes resulted in the incorporation of only a small number of proteins into either vesicles or painted, supported, or suspended bilayers. See Kita-Tokarczyk, K.; Grumelard, J.; Haefele, T.; Meier, W. Polymer 2005, 46, 3540-3563; Ho, D.; Chang, S.; Montemagno, C. D. Nanomedicine 2006, 2, 103-112; and Gonzalez-Perez, A.; Stibius, K. B.; Vissing, T.; Nielsen, C. H.; Mouritsen, 0. G. Langmuir 2009, 25, 10447-10450. While vesicles are excellent vectors for drug delivery, other applications such as sensors, reactive surfaces, drug screening, and water purification would benefit greatly from a planar membrane morphology. See Langer, R. Science 1990, 249, 1527-1533. Also, film rehydration, a technique commonly used to make polymer-protein vesicles, appears to limit the amount of membrane protein that can be incorporated into BCP vesicles, even if high protein concentrations are used. Painted bilayers are excellent tools for studying the function, in particular the conductance, of membrane proteins, but the number of membrane proteins that are incorporated is usually low, and the stability of the membrane can be limited due to the presence of residual solvent. See Mueller, P.; Rudin, D. O.; Tien, H. T.; Wescott, W. C. J. Phys. Chem. 2011, 67, 534-535. Supported and suspended bilayers have also shown low reconstitution of membrane proteins.